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Inspection and related kit for nexin of citrulline chemical fibre

A fibronectin and detection kit technology, which is applied in the field of protein detection, can solve the problem of no disclosed citrullinated fibronectin detection method supporting reagents and the like, and achieve the effects of good practical value, convenient detection tools, and easy popularization and use.

Inactive Publication Date: 2005-12-28
SHANDONG MEDICAL BIO TECH RES CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, after retrieval, there is no published detection method and supporting reagents for citrullinated fibronectin

Method used

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  • Inspection and related kit for nexin of citrulline chemical fibre
  • Inspection and related kit for nexin of citrulline chemical fibre

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Example 1: Determination of Citrullinated Fibronectin in Patient Blood

[0041] (1) Plasma preparation.

[0042] Fresh blood was collected from patients with rheumatoid arthritis, patients with systemic lupus erythematosus, and normal subjects with a blood collection tube containing a final concentration of 3.8% sodium citrate. Centrifuge at 1000X g for 30 minutes. Collect the supernatant. Plasma can be stored at -80°C.

[0043] (2) Solution preparation

[0044] The PBS buffer formula is: 8g / L NaCl, 0.2g / L KCl, 1.15g / L NaHPO 4 and 0.2g / L KH2PO 4 , pH7.5;

[0045] PBS-Tween eluate: add Tween 20 to PBS buffer at a ratio of 1:1000 to prepare, stir and mix.

[0046] (3) Steps:

[0047] A. Dilute the anti-fibrin monoclonal antibody (Abcam company product) 4000 times with 0.05M, pH9.6 carbonic acid / bicarbonate buffer to make its concentration 50ug / ml, add 100ul antibody solution dropwise to In the sample wells of the 96-well ELISA reaction plate, place the plate at 4...

Embodiment 2

[0057] Example 2: Determination of Citrullinated Fibronectin in Patient Blood

[0058] (1) Plasma preparation.

[0059] Fresh blood was collected from patients with rheumatoid arthritis, patients with systemic lupus erythematosus, and normal subjects with a blood collection tube containing a final concentration of 3.8% sodium citrate. Centrifuge at 8000X g for 30 minutes. Collect the supernatant. Plasma can be stored at -80°C.

[0060] (2) Solution preparation

[0061] The PBS buffer formula is: 8g / L NaCl, 0.2g / L KCl, 1.15g / L NaHPO 4 and 0.2g / L KH2PO 4 , pH7.6;

[0062] PBS-Tween eluate: add Tween 20 to PBS buffer at a ratio of 1:1000 to prepare, stir and mix.

[0063] (3) Steps:

[0064] A. Dilute anti-fibrin monoclonal antibody (Abcam company product) 3000 times with 0.05M, pH9.6 carbonic acid / bicarbonate buffer, add 100ul antibody solution dropwise to the sample well of 96-well ELISA reaction plate , place the plate at 4°C for 10-12 hours to cross-link the antibody...

Embodiment 3

[0074] Example 3: Composition of the kit

[0075] 1. Anti-fibronectin pre-coated ELISA plate (bar)

[0076] Take the anti-fibronectin antibody at a ratio of 1:3000, dilute it with PH9.6 carbonate buffer, add 100ul to each well, leave it at room temperature for 4 hours, and wash it three times with PBS containing 0.05% Tween-20. Add 200ul of 2% BSA to the well and set it at 37°C for 2 hours. After drying, blow it dry, seal it in a tin-platinum bag under vacuum, and store it at 4°C.

[0077] 2. Anti-citrulline antibody, conventionally labeled with horseradish peroxidase.

[0078] 3. Product dilution: containing 2% BSA, PBS.

[0079] 4. Washing solution: containing 0.05% Tween-20PBS.

[0080] 5. Color developer: 0.4% TMB.

[0081] 6. Stop solution: 2% H 2 SO 4 .

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Abstract

A kit of carbamyl ornithine fibre connexin consists of pre-coated carrier of antifebrin connexin antibody (ACA), labeled anticarbamyl ornithine antibody (LAOA), diluting liquid, washing solution, color developing agent and terminating liquid. The detecting method includes adding blood plasma in each hole of plate pre-coated with ACA, adding LAOA and adding color developing agent in sequence for reacting, using enzyme mark scale, counter and detector for detecting, comparing detected from sample hole to detected data from control hole and then calculating for result.

Description

technical field [0001] The invention relates to a protein detection method, in particular to a rapid detection method of citrullinated fibronectin and a related kit. technical background [0002] Rheumatoid arthritis is an autoimmune disease. The disease is manifested as chronic polyarthritis, joint swelling and pain, advanced joint tissue damage, and loss of joint function. Rheumatoid arthritis patients contain a variety of autoimmune antibodies, the most important of which is rheumatoid factor. Rheumatoid factor is an immunoglobulin against G-type immunoglobulin. About 70% of patients have rheumatoid factor in their blood. Therefore, the presence or absence of rheumatoid factor is one of the commonly used indicators for clinical diagnosis of rheumatoid arthritis. However, rheumatoid factor is also present in the blood of patients with systemic lupus erythematosus, Sjogren's disease, scleroderma and dermatomyositis, and is often present in the blood of many other chroni...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/543
Inventor 常晓天朱有名韩金祥
Owner SHANDONG MEDICAL BIO TECH RES CENT
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